Characterization and humanization of anti-scorpion venom and anti-TIM-3 nanobodies

• Grant number: 24/03961-6
• Support Opportunities: Research Grants - Visiting Researcher Grant - International
• Start date: June 24, 2024
• End date: August 15, 2024
• Field of knowledge: Biological Sciences - Immunology - Applied Immunology
• Principal Investigator: Daniela Luz Hessel da Cunha
• Grantee: Daniela Luz Hessel da Cunha
• Visiting researcher: Lorena Itati Ibañez
• Visiting researcher institution: Universidad De Buenos Aires/Uba, Argentina
• Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil

Abstract

Recently, the Antibody Development Center at the Butantan Institute began implementing the Center of Nanobody Discovery, led by researchers Daniela Luz, Debora Moretti and Roxane Piazza. This center aims to implement a platform for the selection of therapeutic nanobodies and detection against different antigens, from poisons to infectious and tumor targets with the purpose of generating molecular tools that have promising market characteristics.As part of this implementation, Drs Daniela Luz and Debora Moretti recently established a partnership and research agreements with the National Institute of Agricultural Technology of Argentina (INTA), with Dr Gladys Viviana Parreño as a collaborator to generate immune nanobody libraries from of Llama immunization. Immune libraries were generated against scorpion venom, with the aim of offering new passive immunization tools against poisoning and against the TIM-3 protein, a promising inhibitory receptor, which when blocked has antitumor characteristics. Dr. Parreño is in Brazil via the FAPESP international visiting researcher project (FAPESP n23/07845-8) to implement nanobody selection protocols against these first two targets. The partnership between the two institutions relies on resources from INTA, the Butantan Foundation and regular support from FAPESP, processes 2020/14521-6 (Nanobodies as an alternative therapy for passive immunization) and 2022/04560-0 (Development of platforms for obtaining and characterizing of non-glycosylated recombinant antibodies against the immune checkpoint TIM-3 and evaluation of the role of these antibodies in the reactivation of T lymphocytes in tumors).Given the objectives of this platform, innovative for the Butantan Institute, Dr Parreño introduced Dr Lorena Itati Ibañes from the University of Buenos Aires, Argentina, for her expertise in the characterization and modification of nanobodies. Dr. Lorena Itati Ibañes (UBA/Argentina), researcher at INQUIMAE CONICET, has more than 16 years of experience in the field of Nanobodies. She completed a postdoctoral degree in Dr. Saelens' group (Vlaams Instituut voor Biotechnologie Belgium), in close collaboration with Ablynx, a Sanofi company, a leader in the development of therapeutic Nanobodies. She then returned to Argentina and worked at Inmunova, where she helped establish all the techniques necessary for the production of Nanobodies. She obtained short-term grants to work in New York and Stockholm, where she learned new Nanobody modification techniques. For the last 6 years she has been working on developing a platform for modifying nanobodies for different applications. She created constructs to couple nanobodies to fluorescent proteins, horseradish peroxidase, and alkaline phosphatase. Other modifications include biotinylation of Nanobodies using avitag or Sortase A technologies, addition of Fc tag, among others. Her laboratory routinely produces Nanobodies for the detection and study of different pathogens of animal, human and plant origin.In this proposal, the aim is to establish the characterization of nanobodies generated by immunized Llamas, as well as to modify these tools to better serve therapeutic purposes, such as their humanization. The work plan will then be divided into three fronts: Biochemical characterization, affinity modification and humanization of nanobodies.PV's activity plan therefore includes the completion of the implementation of the nanobody platform at the Butantan Institute, establishing a high affinity ligand, Specific Antigen against scorpion toxin and the TIM-3 protein, which will be important for the establishment this unprecedented platform for the Butantan Institute in addition to establishing solid cooperation between INTA/UBA/Argentina and IB/Brazil, strengthening the partnership between Latin American institutions. (AU)